Corticospinal motor neurons (CSMN) are large pyramidal neurons that reside in layer V of the neocortex, and extend axons to specific segmental targets along the rostro-caudal axis of the spinal cord. The axons of CSMN form the corticospinal tract (CST), which is the major motor output pathway of the motor cortex, and critically controls voluntary movement. Although the anatomical development of CSMN has been welldescribed in many studies in humans and rodents, little is known about the molecular controls over CSMN development, including differentiation and connectivity. Previous studies in our lab (Arlotta et al., Neuron '05; Molyneaux et al., Neuron '05; Ozdinler and Macklis, Nature Neurosci. '06; Molyneaux et al., Nature Rev. Neurosci. '07; Lai et al., Neuron '08; Arlotta et al., J. Neurosci., '08; Joshi et al., '08; Azim et al., '08) have identified developmentally regulated genes that appear to constitute a combinatorial genetic program for control over specification and differentiationof CSMN. These genes are progressively expressed throughout CSMN development, and distinguish CSMN from other closely-related projection neuron subtypes. However, CSMN themselves exhibit a great deal of anatomical heterogeneity: most strikingly, some CSMN extend axons to cervical spinal cord segments, and control forelimb movements, while others extend far more caudally to innervate lumbar segments and control movement of hindlimbs. The genetic basis for this diversity is strongly suggested by the stereotypic organization of CSMN in the motor cortex, which is largely conserved from rodents to primates, and by the precise topographic pattern of connectivity of CSMN axons in the spinal gray matter. Building on these foundations, I aim to investigate the molecular-genetic program for how the diversity and specificity of CSMN segmental connectivity is established during development. I propose to: (1) identify genes specific for cervical- and lumbar-projecting CSMN throughout development, and determine their wild-type expression pattern; (2) characterize the function of select candidate genes differentially expressed between cervical- and lumbar-projecting CSMN through loss- and gain-of-function experiments in vivo. Together, these studies aim to identify critical molecular determinants of the segmental target specificity of CSMN. This work has significant clinical implications, as CSMN are the neuronal population that degenerate in amyotrophic lateral sclerosis (ALS), and play a central role in motor dysfunction following spinal cord injury. Advances in our understanding of the molecular controls over CSMN development can accelerate progress toward protection and repair of cortical circuitry following injury or disease-related neuron death. [unreadable] [unreadable]